The present invention relates to the area of apparatus for the isolation and detection of microbial samples, and particularly to filter apparatus useful with visual microbial detection techniques.
Most of the existing techniques useful in detecting the existence of certain microbial organisms in a given environment, have involved the collection of samples, and the subsequent inoculation of a growth medium therewith, so that a culture may be propagated over a given period of time to determine the presence of the specific microbial organism. Depending upon the microorganism in question, the growth period for such a culture could extend from twenty-four to ninety-six hours and more.
In addition to the growth medium technique discussed above, microbial samples were gathered and stained, and then observed under a microscope. In one technique, a representative liquid sample would be filtered by pressurized application from a syringe to a filter medium. The filter medium would thereafter be dried, and stained. In another technique, concentration of the microbial organism is accomplished by subjecting a liquid sample thereof to centrifugation, after which the concentrated sample is disposed on a microscope slide, and then stained. The last technique comprises the direct application of a small quantity of the microbe-containing liquid on a slide, which is then appropriately stained and viewed under a microscope. All of the foregoing techniques suffer from the shortcomings of being both inaccurate and time consuming. Particularly in the instance where very low concentrations of microorganisms are suspected, these techniques were inadequate and incapable of detecting organism presence.
Recently, a technique has been developed, which is known as the colorimetric method for microbial detection which gives accurate results at very low microbial concentrations. The colorimetric technique comprises the application of a small quantity of an indicator compound to the microbial sample, and positive results are determined by a color reaction caused by the reaction between the indicator compound and the enzymes of the microorganism. This particular technique is accurate to low levels of concentration, however, the present filtration techniques have been inadequate to achieve the desired sample isolation with regularity and dispatch.
The standard equipment currently used for manual filtration such as that conducted in field investigations comprises a disclike filter membrane clamped within a filter holder and between parallel sealing surfaces or gaskets disposed in contact with the opposite broad surfaces of the filter disc. The top half of the filter holder provides a support for the syringe which dispenses the liquid microbial sample. An air space exists between the surface of the filter membrane and the syringe and application of the sample to the filter medium is accomplished by the depression of the plunger on the syringe to release the liquid therefrom.
This manual filter holder suffers from certain shortcomings. Firstly, the gaskets or seals that clampingly retain the filter membrane frequently leak under pressure, with the result that a portion of the liquid sample and its microbial content bypasses the surface of the filter membrane. Secondly, the air space between the filter surface and the syringe frequently causes a fluid build-up in the intermediate space that causes fluid leakage from back-pressure developed in the apparatus. Thus, portions of the sample either escape from the filter holder apparatus or are deposited on the apparatus rather than on the filter medium. Attempts to force the liquid through the filter medium, by the application of compressed air, frequently damages the fragile microbial cells in the sample.